Cerebellar Weight Research Articles

Long-term effects of neonatal ovariectomy on cerebellar development in the rat: a histological and morphometric study

The effects of neonatal ovariectomy on cerebellar development and morphology were investigated in rats ovariectomized (OV) or sham-ovariectomized (SOV) 24 h after birth. At age 70 days rats were sacrificed and the cerebella processed for quantitative light microscopy of matched midline sagittal sections of the vermis. Body, brain and cerebellar weights, as well as total cerebellar area and area of the internal granular layer were significantly greater in the OV rats. The concentration (cells/mm 2) and content (cells/section) of granule cells were significantly greater in each of the 10 vermal lobules of OV rats. The number of Purkinje (P) cells in the OV rats was significantly greater than control values for 6 lobules (L-III, L-V, L-VI b,c, L-VII, L-IX and L-X). P cell number for L-I, L-II and L-VIII was also higher in the OV group, but not significantly. The between group variation for P cell packing density (cells/mm) was significantly different, i.e. packing density for the OV rats was greater than the SOV value. The most consistent and significant difference between the folial patterns of SOV and OV groups was observed in L-VIII, the pyramis. A bilobed L-VIII occurred in 61.11% of SOV rats, but only in 16.67% of OV rats. Overall, neonatal ovariectomy significantly reduced the number of folia. Although the mechanisms are speculative, these data clearly indicate removal of the ovaries 24 h after birth, either directly and/or indirectly has long-term effects on cerebellar development.

Effect of deltamethrin on regional brain polyamines and behaviour in young rats.

The present study examines the mechanism of neurotoxic action of a synthetic pyrethroid formulation deltamethrin in young rats. Newly weaned Wistar Albino male rats received deltamethrin of technical grade at a dose of 7.0 mg/kg body weight/day in corn oil, orally from postnatal day 22 to postnatal day 37. Deltamethrin significantly decreased the wet weight of the hippocampus without much affecting the weight of cerebellum, pons medulla, hypothalamus, frontal cortex and corpus striatum in comparison to respective controls. A significant increase in the activities of mitochondrial monoamine oxidase and microsomal acetylcholinesterase without any effect on microsomal Na+, K(+)-ATPase activity was observed in the brain of experimental animals. Our results further indicate that deltamethrin markedly impaired learning function and significantly increased the spontaneous locomotor activity while aggressive behaviour remained unaffected. An overall enhancement of polyamine levels in hypothalamus and corpus striatum accompanied with an overall decline in pon medulla and cerebellum was also noted. Maximum decrease of spermine and spermidine was registered in hippocampus, while these polyamines showed an increase in frontal cortex. In striatal membranes the binding of 3H-spiperone decreased and 3H-quinuclidinyl benzilate was elevated significantly. Deltamethrin-induced deviations in regional brain polyamine levels may be a possible cause for altered pathophysiology of the neurone.

Impaired neurogenesis by methylazoxymethanol in newborn rats results in transient reduction of ornithine decarboxylase and polyamines in the cerebellum, but not in the olfactory bulbs

Polyamines and the key enzyme for their biosynthesis, ornithine decarboxylase (ODC) play an important role in the control of neuronal proliferation and differentiation. Exposure to agents that interfere with normal cell maturation is expected to result in alteration of neuronal ODC developmental pattern. We have administered to newborn rats, about 6 and 30 hr after birth, 20 mg/kg of methylazoxymethanol acetate (MAM), an agent able to selectively kill dividing cells and we have evaluated ODC activity and polyamine levels in the cerebellum and ODC activity in the olfactory bulbs at various developmental stages starting from postnatal day 4 (PD 4) until PD 28. Cerebellar weight decreased by 22–50% at the different developmental stages in MAM-treated animals. A decline in ODC specific activity was observed at PD 4 and a decrease of putrescine levels at PD 4 and PD 6 in the cerebellum. At PD 10, however, both ODC activity and putrescine level were increased in MAM-treated animals. Spermidine levels were never affected by the treatment, while spermine was significantly decreased at PD 6 and PD 8. These results demonstrate that altered ontogenetic patterns of ODC activity and polyamine levels are the consequence of disturbance of the normal process of brain maturation. No significant differences in specific ODC activity were noticed in the olfactory bulbs of MAM-treated rats. This may be related to the more widespread time-span of neurogenesis in this region, a fact that is also revealed by the higher ODC activity constitutively expressed at times in which neurogenesis has ended in the rest of the brain.

Protective effect of ginseng saponins against impaired brain growth in neonatal rats exposed to ethanol.

This study was performed to determine the active constituents of the root of Panax ginseng C. A. Mayer in the amelioration of ethanol-induced impediment of brain growth in the neonatal stage. To establish an animal model of the brain growth impediment caused by ethanol, ethanol (6 g/kg s.c.) was administered to rat pups on postnatal day 6, which corresponded to the third trimester of pregnancy for humans. Brain weight, especially cerebellar weight, was significantly reduced in the ethanol-exposed pups. In contrast, neither separation from dams nor pentobarbital treatment affected brain weight. A saponin fraction of ginseng extract prevented this ethanol-induced reduction of brain weight. Some ginseng saponins including ginsenosides Rg1, Rb2, Rd, Rf and Re effected stimulated a potent recovery of cerebellum growth in this animal model.

Intragastric intubation of alcohol during postnatal development of rats results in selective cell loss in the cerebellum.

Postnatal alcohol exposure produces reductions in the number of Purkinje cells in the rat cerebellum. The goal of this study was to determine if the method of postnatal alcohol exposure would influence the degree of vulnerability of the Purkinje cells. Previously reported studies from other laboratories have demonstrated cerebellar Purkinje cell count reductions following postnatal alcohol exposure via artificial rearing and vapor inhalation techniques. This study used gastric intubation to administer alcohol (3.6 g ethanol/kg body weight, bid) to male rat pups from postnatal days 4-10. Peak blood alcohol levels were 203 +/- 12.7 mg/dl on postnatal day 6. On postnatal day 10, the animals were perfused, and brain weights were obtained. Body weight was not significantly altered by the postnatal alcohol exposure, yet the wet weights of the cerebral cortex and whole brain were significantly reduced. Although the cerebellar weight was not significantly reduced, the overall number of Purkinje cells measured in the cerebellar vermis was significantly reduced by 24% compared with the isocaloric and normal control groups. The pattern of vulnerability for the individual cerebellar lobules was similar to the previously reported studies, indicating that alcohol's teratogenicity transcends experimental paradigm and is remarkably consistent, when relatively similar blood alcohol profiles are established.

Neurochemical characterization of the alterations in the noradrenergic afferents to the cerebellum of adult rats exposed to X-irradiation at birth.

A single dose of x-irradiation was applied on the cephalic end of newborn rats, and the alterations in the noradrenergic afferents to the cerebellum were studied 180 days later. A net increase in the noradrenaline content of cerebellum was found (122% of nonirradiated controls). The response of noradrenaline content to reserpine injection (0.9 mg/kg, i.p.) was similar in exposed and control rats. Likewise, the 3H release induced by Ro 4-1284 from cerebellar cortex slices labeled with [3H]noradrenaline was unmodified by x-rays, although a mild increase in the spontaneous efflux of 3H was found. The retention of 3H by the slices was reduced in exposed animals (58% of controls). Both the in vitro activity of tyrosine hydroxylase and the accumulation of L-3,4-dihydroxyphenylalanine (L-DOPA) were not significantly different between x-treated rats and controls. In contrast, monoamine oxidase activity was markedly reduced in x-irradiated cerebellum (38% of controls). The x-ray-induced decrease in cerebellar weight (-60%) resulted in marked increases in noradrenaline concentration (223%), tyrosine hydroxylase activity per milligram of protein (206%), and 3H retention (50%). The accumulation of L-DOPA per gram of tissue was also increased at every time considered. These data indicate that x-irradiation at birth produces a cerebellar loss not completely shared by the noradrenergic afferents, and a permanent imbalance between the noradrenergic afferent input and its target cells might eventually result.(ABSTRACT TRUNCATED AT 250 WORDS)

Neurotoxicity of phenytoin administered to newborn mice on developing cerebellum

To examine the neurotoxic effects of phenytoin (PHT) on cerebellar development, we administered 50 mg/kg PHT suspended in sesame oil orally to newborn Jcl:ICR mice once a day during postnatal days 2–14 and determined plasma PHT concentrations at designated intervals during the administration period. In the treated group, walking reflex and negative geotaxis were poorly developed on postnatal day 14. Pyknotic cells in the external granular layer (EGL) significantly increased and were prominent in the vermis area compared with controls on postnatal day 14. Plasma PHT levels were 34–36 μg/ml on the 3rd day of PHT treatment and approached a steady-state situation. Total brain weight, size of the cerebellum, and cerebellar weight were significantly reduced in the treated group on postnatal day 56. Accordingly, oral administration of PHT in the neonatal period induced neurotoxic damage on the developing cerebellum.

Effect of prenatal and postnatal exposure to ethanol on rat central nervous system gangliosides and glycosidases

We investigated the effect of maternal alcohol consumption on cell number, gangliosides and ganglioside catabolizing enzymes in the central nervous system (CNS) of the offspring. Virgin female rats of the Charles Foster strain were given 15% (v/v) ethanol in drinking water one month prior to conception and during gestation and lactation. At 21 days postnatal age, the offspring were sacrificed and the brains were separated into cerebrum, cerebellum and brain stem to investigate possible regional variations. Compared to controls, wet weight of cerebrum, cerebellum and brain stem, and of spinal cord was decreased in the pups exposed to alcohol. DNA and protein contents were also found to be lowered in all the CNS regions of the pups exposed to alcohol. Conversely, maternal alcohol consumption was found to increase the concentration and the content of total ganglioside N-acetyl-neuraminic (NANA) in CNS of the pups. In addition, alcohol treatment was found to induce alterations in the proportions of individual ganglioside fractions. Interestingly, these alterations are somewhat different than those observed in the neonatal brain and spinal cord of the pups subjected to prenatal alcohol exposure. The alterations in the proportions of ganglioside fractions were shown to be region-specific. Maternal alcohol consumption resulted in decreased activities of sialidase, beta-galactosidase, beta-glucosidase and beta-hexosaminidase. The results suggest that the alcohol-associated increases in ganglioside concentration may be at least partly due to the decreased activities of ganglioside catabolizing enzymes.

Accurate prediction of Purkinje cell number from cerebellar weight can be achieved with the fractionator

Purkinje cell nucleoli are used as counting units in order to obtain unbiased (fractionator) estimates of the number, N, of Purkinje neurons in adult mammalian cerebella of known weight, W. Regression analysis is then employed to establish the nature of the relationship between logN and logW. The linear regression equation defines an allometric relation that is employed to predict number in cerebella of known weight from other mammals. Predicted numbers are tested against empirical estimates. For 19 cerebella ranging in weight from 0.2 g (rat) to 113 g (human), the allometric relation between Purkinje cell number and organ weight was determined. By using this relation, the mean complement in three rabbit cerebella (average weight, 0.87 g) is predicted to be 0.63 million. This figure is confirmed by fractionator estimates made on the same three brains. The cat cerebellum should contain about 1.5-2.0 million Purkinje cells. An estimate of 1.2-1.3 million cells is to be found in the literature. Including rabbit cerebella in a refined equation yields the following relation: N = 686000W(0.695). With this refined equation, further predictions are made about the numbers likely to be found in the cerebella of the dog, goat, pig, ox, and horse. The numbers predicted for these animals must await experimental verification, but they are entirely consistent with previous suggestions that neuronal packing densities decrease with increasing brain size.

Growth rates and patterns of organs and tissues in the bovine fetus

Ninety-two purebred Jersey fetuses from clinically normal dams derived from a variety of sources but unexposed to experimental infection, were killed between 80 and 260 days after conception and dissected to provide basic parameters of a normal population. Organs and tissues were weighed and measured and the equations for the regression of the values on fetal age evaluated. Growth followed a sigmoid curve with rapid changes in growth rate between 140 and 170 days' gestation. While a few variables increased their growth rate over the late fetal period, the majority maintained a low even rate of growth. Thyroid and cerebellum weights showed a marked reduction in growth rate at this time. Long bone length and crown-anus length were the most predictable parameters for a given gestational age.

Long-term deficits in cerebellar growth and rotarod performance of rats following “binge-like” alcohol exposure during the neonatal brain growth spurt

The cerebellum is vulnerable to growth restriction and neuronal depletion induced by alcohol exposure during the brain growth spurt of neonatal rats. This study examined whether neonatal alcohol exposure permanently restricted brain growth and induced motor performance deficits in adults. Two groups of rats were given 4.5 g/kg of alcohol per day during postnatal days 4 through 9, using artificial-rearing procedures. One group was given the alcohol as a 10.2% (v/v) solution in two of the 12 daily feedings, producing peak BACs of 361 mg/dl. The second group was given the alcohol as a 5.1% (v/v) solution in four of the feedings, producing peak BACs of 187 mg/dl. Controls included an artificially reared group and a normally reared group. All rats were tested on a rotarod at approximately 405 days of age, then perfused 1–2 weeks later. The 10.2% group was significantly impaired in acquiring the task and had significant reductions in whole brain and cerebellar weight, compared to controls. The 5.1% treatment also significantly restricted whole brain and cerebellar growth, and rotarod performance of that group was intermediate between the control groups and the 10.2% group. The cerebellar reductions and deficits in motor performance in adulthood demonstrate permanent structural and functional consequences of binge-like alcohol exposure during the brain growth spurt.

Effect of Protein Malnutrition on Glycoprotein, Protein and Lipid Synthesis in the Rat Cerebellum during the Period of Brain Growth Spurt

Female Wistar rats were fed a normal-protein diet (25% casein) or a low-protein diet (8% casein) during pregnancy and lactation. The two diets were isocaloric and contained appropriate amounts of mineral salts and vitamins. Pups from dams submitted to the low-protein diet had a lower body weight than normally fed controls as early as on the day of birth, but a difference in cerebellar weight between the two groups was observed only on the 15th postnatal day. Malnutrition had no effect on cerebellar protein concentration, which increased with age in both groups. The cerebellar DNA concentration was higher at 7 and 15 days of age in normally fed rats than in malnourished rats, whereas at 21 days of age it was higher in the malnourished animals. [U-14C]Leucine and [2-3H]mannose incorporation into proteins and lipid synthesis from acetyl coenzyme A (CoA) derived from [U-14C]leucine markedly decreased with age in the cerebellum of rats fed both diets. [2-3H]Mannose incorporation into cerebellar glycoproteins was greater in malnourished rats during the period of brain growth spurt than in normally fed rats at all ages studied. Prenatal and postnatal protein malnutrition had no effect on [U-14C]leucine incorporation into cerebellar proteins or on cerebellar lipid synthesis from acetyl-CoA derived from [U-14C]leucine during the period of brain growth spurt.

Increased activity of tyrosine hydroxylase in the cerebellum of the X-irradiated dystonic rat

The exposure of the cephalic end of rats to repeated doses of X-irradiation (150 rad) immediately after birth induces a long-term increase in the noradrenaline (NA) content of cerebellum (CE) (+ 37.8%), and a decrease in cerebellar weight (65.2% of controls), which results in an increased NA concentration (+ 109%). This increase in the neurotransmitter level is accompanied by a dystonic syndrome and histological abnormalities: Purkinje cells (the target cells for NA afferents to CE) fail to arrange in a characteristic monolayer, and their primary dendritic tree appears randomly oriented. The injection of reserpine 0.9 and 1.2 mg/kg ip to adult rats for 18 h depletes cerebellar NA content in both controls (15.7 +/- 4 ng/CE and 2.8 +/- 1.5 ng/CE, respectively) and X-irradiated rats (17.1 +/- 1 ng/CE and 8.3 +/- 2 ng/CE, respectively). The activity of tyrosine hydroxylase (TH) in CE of adult rats, measured by an in vitro assay, is significantly increased in neonatally X-irradiated animals when compared to age-matched controls (16.4 +/- 1.4 vs 6.32 +/- 0.6 nmol CO2/h/mg prot., p less than 0.01). As observed for NA levels, a net increase in TH activity induced by the ionizing radiation is also measured: 308.9 +/- 23.8 vs 408.2 +/- 21.5 nmol CO2/h/CE, p less than 0.01 (controls and X-treated, respectively). These results suggest that X-irradiation at birth may induce an abnormal sprouting of noradrenergic afferents to CE. The possibility that these changes represent a response of the NA system to the dystonic syndrome is discussed.

Cerebellar ganglioside abnormalities in pcd mutant mice

The distribution of cerebellar gangliosides was studied in Purkinje cell degeneration (pcd/pcd) mutant mice at postnatal days 25, 30, 50, and 150. These mutants lose the majority of Purkinje cells between 18 and 50 days of age. A reactive gliosis accompanies Purkinje cell loss and a partial loss of granule cells occurs in pcd/pcd mice older than p50. Purkinje cell loss is associated with significant reductions in cerebellar weight and ganglioside concentration. This neuronal loss was also developmentally correlated with reductions of gangliosides (GT1a/LD1 and GT1b and with elevations of GD3. These results agree with previous findings in other cerebellar mutants that GT1a/LD1 and GT1b are concentrated in Purkinje cells and that GD3 is enriched in reactive glial cells. A slight, but significant, reduction in GD1a concentration occurred only in older pcd/pcd mice, consistent with previous findings in weaver and staggerer mice that GD1a is enriched in mature granule cells. The findings with pcd/pcd and other neurological mutants indicate that certain gangliosides can serve as cell-surface markers for monitoring changes in cerebellar cytoarchitecture that accompany development or disease.

Artificial rearing of rat pups using rat milk

The contribution of diet and surgery to the brain weight deficits observed in artificially reared rats was investigated. Four day old Long Evans rat pups were assigned to an artificially reared (AR) or mother reared (MR) group. AR pups were encannulated and fed either rat milk (AR-MOM) or replacement formula (AR-MES). MR pups received a sham encannulation (MR-SHAM) or no surgery (MR-CONT) before being returned to their dam for rearing. On day 7 all the animals were killed. Brain weights and visceral organ weights were obtained. There was no significant difference between the MR groups on any measure except stomach weights. AR-MOM pups had larger visceral organ weights than pups in the other groups. AR-MOM and AR-MES pups had similar whole brain weights, smaller than those of the MR pups. However, the cerebellar weights, and to a lesser extent, brainstem weights, showed improvements in the AR-MOM group, over the AR-MES group. Neither the effect of surgery nor of diet alone can account for the organ weight differences that have been described in AR rats. The possibility that normal growth may be primarily dependent on diet at one stage of development, with other factors gaining importance at later stages is discussed.

Structural and neurochemical effects in mouse cerebellum following neonatal methylazoxymethanol and 6-hydroxydopamine treatment

The effects of neonatal treatment with the antimitotic agent methylazoxymethanol and the catecholamme neurotoxin 6-hydroxydopamine on cerebellar morphology and monoamine innervation in the N.M.R.I. mouse has been studied. Methylazoxymethanol (25 mg/kg s.c.) treatment induced a cerebellar weight reduction of 40% as observed in the adult stage, while other CNS regions analysed were unaffected. An obvious atrophy of the cerebellar cortex was found, with an irregular distribution of the Purkinje cells, while Bergmann glia fibers deviated from their normal radial configuration and showed a tendency to form clusters. A 65% increase of tyrosine hydroxylase immunoreactive fiber density was found in the cerebellar cortex and 3H-5-hydroxytryptamine in vitro synaptosomal uptake was increased by 55%. Noradrenaline and 5-hydroxytryptamine concentrations in the cerebellum increased by 50 and 30%, respectively, whereas the total content of both neurotransmitters in cerebellum was approximately unchanged after methylazoxymethanol treatment. A significant reduction in total cerebellar in vitro binding of 3H-WB-4101 and 3H-dihydroalprenolol was also found, indicating compensatory receptor alterations following methylazoxymethanol treatment. The effect of combined treatment of methylazoxymethanol and the neurotoxin 6-hydroxydopamine (50 mg/kg s.c., day 1) showed a very pronounced reduction of noradrenaline concentration in cortex cerebri, while the noradrenaline concentration in cerebellum was increased by 185% and the tyrosine hydroxylase immunoreactive fiber density by 125%, indicating an additional relative hyperinnervation of cerebellar noradrenaline fiber due to a “pruning effect” of the 6-hydroxydopamine treatment. The results imply a relatively rigid development of terminal arborization of central nervous system monoamine neurons, relatively independent of neuronal and glial arrangement in the target area.

On the number of Purkinje cells in the human cerebellum: unbiased estimates obtained by using the "fractionator".

Stereological estimates of the numbers of Purkinje cell nucleoli in human cerebellar cortex have been obtained from systematic random samples of tissue by using the fractionator. The estimates are unbiased by fixation, section thickness, or sampling errors and are independent of any assumptions about cell shape, size, or spatial orientation. Twelve brains from aged subjects of both sexes were examined. The average complement of nucleoli in four female brains (age range 71-93 years) amounted to 14.8 millions (with an observed coefficient of variation between subjects of 29%). For three male brains (76-91 years), the corresponding estimates were 15.7 millions (10%). No significant sex differences were found for these small samples. Five brains of unknown sex and age yielded values of 15.8 millions (18%). For the twelve brains examined, the total number of Purkinje cell nucleoli per cerebellum was found to be 15.4 millions (19%). Estimated numbers showed a significant positive correlation with cerebellar weights. The number of nucleoli in an individual cerebellum was obtained with high precision in as short a time as 4 hours.

Relative cerebellar weight: A potential indicator of developmental neurotoxicity

Overt malformations do not always accompany behavioral changes resulting from perinatal exposure to certain drugs. However, the central nervous system (CNS) is the anatomical substrate for behavior, and functional defects may be accompanied by more subtle, structural alterations of the brain. The purpose of this study was to determine if changes in the weights of certain brain regions occur in rats exposed prenatally and/or perinatally to propylthiouracil (PTU) which retards functional development of the brain. Pregnant rats were dosed with PTU during gestation and/or lactation, and on postnatal day 28, auditory startle responses were measured to determine if PTU altered functional development. The brains of all pups were then dissected into 10 separate regions, dried and individually weighed. Brain weights were expressed in absolute and relative (to total brain weight) terms. Pups that were exposed to PTU from days 10 to 21 of gestation grew normally, but their startle responses to auditory stimuli were significantly different ( p<0.05) from controls. Thus, for the purpose of this experiment, PTU acted as a developmental neurotoxicant because it altered performance on a test of neuromuscular function without being overtly teratogenic. The weights of most brain regions in PTU-treated rats were statistically comparable to controls. However, relative cerebellar weight was significantly ( p<0.05) different. Therefore, these data suggested that relative cerebellar weight might be used to predict functional defects that appear during development following prenatal or perinatal exposure to certain neurotoxicants. Although cerebellar weight and auditory startle responses were altered by PTU, this study does not establish a causal relationship between the anatomical and functional changes that occurred. However, the anatomical correlation of altered startle responses suggests that relative cerebellar weight may be a useful measure of developmental neurotoxicity. Since neurogenesis in the rat cerebellum begins early in gestation and continues for several weeks after birth, it is vulnerable to perturbation by drugs for an extended time during development Thus cerebellar weight, which is easily monitored, may be an anatomical predictor of functional development that can be used to complement findings from more conventional performance tests for developmental neurotoxicity.

Aspirin augments alcohol in restricting brain growth in the neonatal rat

The purpose of this study was to determine whether the microencephaly in rats resulting from early postnatal alcohol exposure is altered by a concurrent administration of aspirin. Neonatal rats were artificially reared from postnatal day 4 to postnatal day 10, a period of the brain growth spurt in the rat that is similar to the third trimester of human brain development. The alcohol-treated groups received 6.6 g/kg/day of ethanol and either 0.0, 12.5, 25.0 or 50.0 mg/kg/day of aspirin in a milk solution. Control groups received either 0.0 (gastrostomy control), 12.5, 25.0 or 50.0 mg/kg/day of aspirin in a milk solution free of alcohol. Brainstem, cerebellum and total brain weights were measured on postnatal day 10. Alcohol alone significantly reduced the mean total brain weight, cerebellum and brainstem weight by 19.8%, 23.1% and 12.2%, respectively, relative to gastrostomy controls. A significant interaction between ethanol and aspirin was observed for total brain weight. The mean total brain weight of the group receiving both alcohol and 50 mg/kg/day aspirin was significantly lower than all other experimental groups and was reduced 29.5%, relative to gastrostomy controls. The highest dose of aspirin alone significantly reduced cerebellar weight, relative to gastrostomy controls but had no effect on brainstem or total brain weight.

Developmental Profile of Kappa, Mu and Delta Opioid Receptors in the Rat and Guinea Pig Cerebellum

The phenotypic expression of the three opioid receptors kappa, mu and delta was determined in the rat and guinea pig cerebellum during ontogeny. In both species, the increase in cerebellum weight was accompanied with a gradual increase in the total number of opioid receptors. At the same time, however, the receptor concentration (femtomoles per milligram protein) fluctuated, and finally declined by 3.3- and 2.0-fold in the adult rat and guinea pig cerebella, respectively. In the guinea pig, kappa receptors constituted about 80% of the receptors at the earliest developmental stage tested, day 20 of the embryo, but then there was a sharp decrease in this value, resulting from an increase in both mu and delta receptors. This decrease was, however, transient, and the percentage of the kappa receptors increased again to reach 77% of the receptors in the adult cerebella. A similar transient increase in the percentage of mu and delta receptors was observed during the development of the rat cerebellum, although the receptor density in this species was 1/4-1/8 that of the guinea pig. Cultures of aggregating guinea pig cerebellar cells were used to further study the factors regulating the expression of the three opioid receptors in this brain region. The possibility that the development in vivo of cells expressing different opioid receptors is associated with the ontogeny of the endogenous opioid peptides is discussed.